A blood test to identify cognitively unimpaired individuals at risk for Alzheimer's disease (AD) and the emergence of AD in Down syndrome (DS) is desperately needed. Here, we propose experiments to address this unmet need. Specifically, we will measure (in plasma) levels of proteins (tau and A?) implicated in the etiology of AD, and naturally occurring autoantibodies (NAbs) against these proteins. In blood tau and A? are present in distinct pools, e.g. free floating, bound to other proteins, and inside exosomes. Numerous prior studies have measured A? in plasma, but few took account of the distinct pools and many were confounded by imperfect assays and/or patient specimens. Only a handful of studies have looked at tau in blood, none accounted for the molecular heterogeneity of tau or its occurrence in different pools. In contrast, we will be careful to use assays capable of detecting distinct forms of tau and to assess the contributions of different pools and how they may change with disease. Since tau is present in blood at very low levels and measurement of A? in plasma requires dilution to overcome matrix effects, we will use state-of-the-art in-house ultra-sensitive assays developed using the Simoa (tau) and Erenna (A?) platforms. NAbs-bound tau and A? will be liberated from antigen-antibody complexes and then measured, and free NAbs will be detected by quantifying binding to plate-immobilized antigen. Great care will be taken to include internal standards so as to monitor the sensitivity and reproducibility of our assays. Most prior studies evaluated analytes at only a single time point and there have been few longitudinal studies. Considering the long duration of both the pre-clinical and clinical phases of AD, and how distinct forms of tau, A? and NAbs may change during these protracted periods, it is not surprising that cross-sectional studies have, thus far, yielded conflicting and variable results. Mindful of these pitfalls, we propose for the first time to measure anti-tau and anti-A? NAbs and their cognate antigens in the same plasma samples using specimens from carefully characterized study subjects. To gain insight on how tau, A? and NAbs change throughout the course of the disease, we will apply our well- characterized assays to plasma samples from 3 distinct cohorts. These will include samples that have been collected prospectively before and just after clinical onset from individuals of whom a detailed set of clinical, genetic, and histopathology data exist, and plasma from 0-65 years old DS subjects. DS is the most common genetic cause of early-onset AD and most DS adults become demented before the age of 60. Thus, studying samples from different-aged DS subjects provides a window on different stages of AD, and should also identify the optimal therapeutic interval to treat AD in DS. Combining this systematic approach with careful statistical analysis we expect to identify one, or possibly a selection of analytes, that can: (i) accurately detect subjects at risk of developing AD, and (ii) determine an appropriate age range in which to treat AD in DS.